There are noteworthy and fundamental similarities between inframammalian and mammalian vertebrates in their cellular responses to spinal cord injury, even though functional regeneration occurs only in some of the former species and never in mammals. The progressive tissue necrosis that is characteristic of spinal injury in mammals may be largely responsible for aborting incipient neuroregenerative efforts (i2). This possibility is supported by experiments which demonstrate extensive growth of CNS axons into non-necrotizing PNS environments or into spinal lesions whre necrosis is prevented by maintaining the animals in hibernation. We recently tested this hypothesis by injuring the spinal cord of rats in such a way as to produce axotomy with minimal post-traumatic tissue necrosis. We observed that neurites did indeed grow into the lesion site (59) and that superfusion of this region with cytosine arabinoside (an antimitotic agent) or triethanolamine (a proton scavenger) markedly enhanced axonal growth into the injured tissue. To make further progress in this in-vivo analysis of spinal injury, we must utilize methods that offer greater quantitative precision and cytological specificity. We therefore propose to adapt a quantitative immunochemical procedure to the assay of selected antigens in tissue sections. Adjacent serial frozen sections of unfixed rat spinal cord will then be examined in sequence for neurofilament protein, glial fibrillary acidic protein, myelin basic protein, laminin, and fibronectin by immunocytochemistry and by quantitative immunochemistry. Conventional histological staining procedures will provide a basis for comparison. In this way we will be able to analyze the entire extent of a spinal cord lesion for localization and for content of specific constituents. This analysis will be performed at various postoperative stages after types of injuries where necrosis is minimal (spinal crush) or extensive (spinal transection) in order to resolve the following fundamental issues: (i) What are the interactions between growing neurites and the cellular and subcellular features of their environment that initiate, maintain, and orient neuritic growth? (ii) What are the sources, trajectories, and destinations of the neurites which grow into a non-necrotizing lesion? (iii) Will reduction of necrosis after spinal transection result in enhanced neuritic growth into the lesion?